DE3785726T2 - METHOD FOR CONTROLLED DEPOLYMERIZATION OF POLYSACCHARIDES. - Google Patents
METHOD FOR CONTROLLED DEPOLYMERIZATION OF POLYSACCHARIDES.Info
- Publication number
- DE3785726T2 DE3785726T2 DE8787116225T DE3785726T DE3785726T2 DE 3785726 T2 DE3785726 T2 DE 3785726T2 DE 8787116225 T DE8787116225 T DE 8787116225T DE 3785726 T DE3785726 T DE 3785726T DE 3785726 T2 DE3785726 T2 DE 3785726T2
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- Germany
- Prior art keywords
- heparin
- reaction
- polysaccharides
- polysaccharide
- depolymerization
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- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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- 238000000034 method Methods 0.000 title claims description 15
- 150000004676 glycans Chemical class 0.000 title claims description 13
- 229920001282 polysaccharide Polymers 0.000 title claims description 13
- 239000005017 polysaccharide Substances 0.000 title claims description 13
- 229920000669 heparin Polymers 0.000 claims description 20
- 229960002897 heparin Drugs 0.000 claims description 19
- HTTJABKRGRZYRN-UHFFFAOYSA-N Heparin Chemical compound OC1C(NC(=O)C)C(O)OC(COS(O)(=O)=O)C1OC1C(OS(O)(=O)=O)C(O)C(OC2C(C(OS(O)(=O)=O)C(OC3C(C(O)C(O)C(O3)C(O)=O)OS(O)(=O)=O)C(CO)O2)NS(O)(=O)=O)C(C(O)=O)O1 HTTJABKRGRZYRN-UHFFFAOYSA-N 0.000 claims description 12
- 238000006243 chemical reaction Methods 0.000 claims description 10
- 229920000045 Dermatan sulfate Polymers 0.000 claims description 4
- 229940051593 dermatan sulfate Drugs 0.000 claims description 4
- AVJBPWGFOQAPRH-FWMKGIEWSA-L dermatan sulfate Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@H](OS([O-])(=O)=O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@H](C([O-])=O)O1 AVJBPWGFOQAPRH-FWMKGIEWSA-L 0.000 claims description 4
- 239000002904 solvent Substances 0.000 claims description 4
- 239000007864 aqueous solution Substances 0.000 claims description 3
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 claims description 3
- QWPPOHNGKGFGJK-UHFFFAOYSA-N hypochlorous acid Chemical compound ClO QWPPOHNGKGFGJK-UHFFFAOYSA-N 0.000 claims description 3
- 239000008194 pharmaceutical composition Substances 0.000 claims description 3
- 229920002971 Heparan sulfate Polymers 0.000 claims description 2
- 238000001311 chemical methods and process Methods 0.000 claims description 2
- 239000003638 chemical reducing agent Substances 0.000 claims description 2
- 239000000243 solution Substances 0.000 description 9
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 6
- 229920002683 Glycosaminoglycan Polymers 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- DWAQJAXMDSEUJJ-UHFFFAOYSA-M Sodium bisulfite Chemical compound [Na+].OS([O-])=O DWAQJAXMDSEUJJ-UHFFFAOYSA-M 0.000 description 3
- WQYVRQLZKVEZGA-UHFFFAOYSA-N hypochlorite Chemical compound Cl[O-] WQYVRQLZKVEZGA-UHFFFAOYSA-N 0.000 description 3
- 235000010267 sodium hydrogen sulphite Nutrition 0.000 description 3
- 238000000196 viscometry Methods 0.000 description 3
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 2
- 102000004411 Antithrombin III Human genes 0.000 description 2
- 108090000935 Antithrombin III Proteins 0.000 description 2
- 229960005348 antithrombin iii Drugs 0.000 description 2
- 239000003054 catalyst Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 230000003647 oxidation Effects 0.000 description 2
- 238000007254 oxidation reaction Methods 0.000 description 2
- 238000007086 side reaction Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- KZBUYRJDOAKODT-UHFFFAOYSA-N Chlorine Chemical compound ClCl KZBUYRJDOAKODT-UHFFFAOYSA-N 0.000 description 1
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 description 1
- KRKNYBCHXYNGOX-UHFFFAOYSA-K Citrate Chemical compound [O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O KRKNYBCHXYNGOX-UHFFFAOYSA-K 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 208000032843 Hemorrhage Diseases 0.000 description 1
- IOVCWXUNBOPUCH-UHFFFAOYSA-N Nitrous acid Chemical compound ON=O IOVCWXUNBOPUCH-UHFFFAOYSA-N 0.000 description 1
- 229940124158 Protease/peptidase inhibitor Drugs 0.000 description 1
- 239000005708 Sodium hypochlorite Substances 0.000 description 1
- 206010047249 Venous thrombosis Diseases 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 238000005903 acid hydrolysis reaction Methods 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 238000005904 alkaline hydrolysis reaction Methods 0.000 description 1
- 230000002785 anti-thrombosis Effects 0.000 description 1
- 239000003146 anticoagulant agent Substances 0.000 description 1
- 229940127219 anticoagulant drug Drugs 0.000 description 1
- 229960004676 antithrombotic agent Drugs 0.000 description 1
- 238000007068 beta-elimination reaction Methods 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 238000004061 bleaching Methods 0.000 description 1
- 230000023555 blood coagulation Effects 0.000 description 1
- 238000010504 bond cleavage reaction Methods 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 239000013043 chemical agent Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000005660 chlorination reaction Methods 0.000 description 1
- 239000000460 chlorine Substances 0.000 description 1
- 229910052801 chlorine Inorganic materials 0.000 description 1
- 239000007979 citrate buffer Substances 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- 150000002016 disaccharides Chemical group 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 238000005194 fractionation Methods 0.000 description 1
- 229910001385 heavy metal Inorganic materials 0.000 description 1
- ZFGMDIBRIDKWMY-PASTXAENSA-N heparin Chemical compound CC(O)=N[C@@H]1[C@@H](O)[C@H](O)[C@@H](COS(O)(=O)=O)O[C@@H]1O[C@@H]1[C@@H](C(O)=O)O[C@@H](O[C@H]2[C@@H]([C@@H](OS(O)(=O)=O)[C@@H](O[C@@H]3[C@@H](OC(O)[C@H](OS(O)(=O)=O)[C@H]3O)C(O)=O)O[C@@H]2O)CS(O)(=O)=O)[C@H](O)[C@H]1O ZFGMDIBRIDKWMY-PASTXAENSA-N 0.000 description 1
- 239000002565 heparin fraction Substances 0.000 description 1
- 229960001008 heparin sodium Drugs 0.000 description 1
- 238000001802 infusion Methods 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 238000011866 long-term treatment Methods 0.000 description 1
- 229940127215 low-molecular weight heparin Drugs 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 239000002808 molecular sieve Substances 0.000 description 1
- 239000000137 peptide hydrolase inhibitor Substances 0.000 description 1
- 150000002978 peroxides Chemical class 0.000 description 1
- 150000004965 peroxy acids Chemical class 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 238000000053 physical method Methods 0.000 description 1
- 230000003389 potentiating effect Effects 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 238000007348 radical reaction Methods 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 230000007017 scission Effects 0.000 description 1
- URGAHOPLAPQHLN-UHFFFAOYSA-N sodium aluminosilicate Chemical compound [Na+].[Al+3].[O-][Si]([O-])=O.[O-][Si]([O-])=O URGAHOPLAPQHLN-UHFFFAOYSA-N 0.000 description 1
- 229940079827 sodium hydrogen sulfite Drugs 0.000 description 1
- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical compound [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 description 1
- 208000010110 spontaneous platelet aggregation Diseases 0.000 description 1
- 206010043554 thrombocytopenia Diseases 0.000 description 1
- 238000011282 treatment Methods 0.000 description 1
- 238000000108 ultra-filtration Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
- C08B37/0063—Glycosaminoglycans or mucopolysaccharides, e.g. keratan sulfate; Derivatives thereof, e.g. fucoidan
- C08B37/0075—Heparin; Heparan sulfate; Derivatives thereof, e.g. heparosan; Purification or extraction methods thereof
- C08B37/0078—Degradation products
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Molecular Biology (AREA)
- Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Materials Engineering (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Medicinal Chemistry (AREA)
- Polymers & Plastics (AREA)
- Organic Chemistry (AREA)
- Polysaccharides And Polysaccharide Derivatives (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Description
Die vorliegende Erfindung betrifft ein neues Verfahren für eine kontrollierte chemische Depolymerisation von natürlich vorkommenden linearen Polysacchariden, insbesondere von Glucosaminoglykanen (GAG).The present invention relates to a new process for a controlled chemical depolymerization of naturally occurring linear polysaccharides, in particular glucosaminoglycans (GAG).
Diese Erfindung umfaßt die gemäß dem Verfahren erhaltenen, depolymerisierten Heparine und pharmazeutische Zusammensetzungen, die solche depolymerisierten Heparine enthalten.This invention encompasses the depolymerized heparins obtained according to the process and pharmaceutical compositions containing such depolymerized heparins.
Eine kontrollierte Depolymerisation einiger natürlicher GAGs, wie Heparin, Heparansulfat (Hs), Dermatansulfat (DS), ist eine reizvolle, aber, in Praxis, schwierige Aufgabe.Controlled depolymerization of some natural GAGs, such as heparin, heparan sulfate (Hs), dermatan sulfate (DS), is an attractive but, in practice, difficult task.
Interesse entsteht aus einigen experimentellen Beobachtungen, wonach Bioverfügbarkeit und biologische Wirkungen einiger GAGs, oral oder parenteral in experimentellen Modellen oder an Menschen verabreicht, von dem mittleren Molekulargewicht (MG) abhängen. Zum Beispiel ist kommerzielles Heparin ein gut bekanntes Antikoagulans aufgrund seiner aktivierenden Eigenschaften als ein Cofaktor auf einen wirksamen Protease-Inhibitor, Antithrombin III (At-III). Heparin ist ein sulfatiertes GAG mit einem = 12000 D, entsprechend ungefähr 20 Disaccharid-Einheiten, und bei der Blutgerinnung inhibiert es 5 Faktoren, nämlich F XII, F XI, F IX, F X, F II. Kürzlich wurde entdeckt, daß Heparinfraktionen mit einem = 4000 - 6000 D (Niedermolekulargewicht-Heparine, NMGH) die Inhibierung von einigen Faktoren, jedoch nicht von anderen beeinflußen und insbesondere von einigen Schlüsselfaktoren beim Venenthrombosebeginn, wie F X. Folglich konnte NMGH als wirksame Antithrombotika mit geringerem Hämorrhagie-Risiko als kommerzielles Heparin angesehen werden.Interest arises from some experimental observations that bioavailability and biological effects of some GAGs, administered orally or parenterally in experimental models or in humans, depend on the average molecular weight (MW). For example, commercial heparin is a well-known anticoagulant due to its activating properties as a cofactor to a potent protease inhibitor, antithrombin III (At-III). Heparin is a sulfated GAG with a = 12000 D, corresponding to approximately 20 disaccharide units, and in blood coagulation it inhibits 5 factors, namely F XII, F XI, F IX, FX, F II. Recently it was discovered that heparin fractions with a = 4000 - 6000 D (low molecular weight heparins, NMGH) affect the inhibition of some factors but not others, and in particular of some key factors in the onset of venous thrombosis, such as F X. Consequently, NMGH could be considered as an effective Antithrombotics with a lower risk of hemorrhage than commercial heparin.
Es ist auch gefunden worden, daß die Halbwertszeit für NMGH im Blutstrom länger ist als für normales Heparin. Es kann daher eine Langzeitbehandlung durchgeführt werden, die Tropfinfusionen und Rebound-Phänomene vermeidet.It has also been found that the half-life for NMGH in the bloodstream is longer than for normal heparin. Long-term treatment can therefore be carried out, avoiding drip infusions and rebound phenomena.
Ein weiterer Vorteil für NMGH ist das Fehlen einer Einwirkung auf die Blutplättchenaggregation, ein unerwünschter Effekt, der in der Vergangenheit schwere Thrombozytopenie bei einigen Patienten während der Heparinverabreichung hervorrief.Another advantage of NMGH is the lack of interference with platelet aggregation, an undesirable effect that has historically caused severe thrombocytopenia in some patients during heparin administration.
In der gegenwärtigen Praxis ist die kontrollierte Depolymerisation von Heparin und einigen anderen GAGs aus vielen Gründen eine schwierige Aufgabe:In current practice, the controlled depolymerization of heparin and some other GAGs is a difficult task for many reasons:
a) Chemische Mittel wie sie zur Zeit verwendet werden (salpetrige Säure, ß-Eliminierung, Oxidation, saure oder alkalische Hydrolyse) modifizieren die molekulare Struktur um die angegriffenen Bindungen herum oder in anderen Regionen, die der Ort für die Aktivität sind, und verursachen dadurch Artefakte.a) Chemical agents currently used (nitrous acid, ß-elimination, oxidation, acid or alkaline hydrolysis) modify the molecular structure around the attacked bonds or in other regions that are the site of activity, thereby causing artifacts.
b) Enzymatische Spaltung ist schwierig zu handhaben, wenn die Depolymerisation bei einem im voraus gewählten Punkt gestoppt werden muß. Weiterhin werden Heparin, HS, DS nicht leicht durch üblicherweise erhältliche Enzyme angegriffen.b) Enzymatic cleavage is difficult to handle if the depolymerization must be stopped at a preselected point. Furthermore, heparin, HS, DS are not easily attacked by commonly available enzymes.
c) Fraktionierung durch physikalische oder chemischphysikalische Methoden, wie z.B. Molekularsieben, Ultrafiltration oder selektive Fällung, ist nicht geeignet für eine großtechnische, wirtschaftliche Herstellung.c) Fractionation by physical or chemical-physical methods, such as molecular sieves, ultrafiltration or selective precipitation, is not suitable for large-scale, economical production.
Eine kontrollierte chemische Depolymerisation von natürlichen Polysacchariden wie Heparinen durch eine Radikalreaktion in Gegenwart eines Metallkatalysators und eines Peroxids oder einer Persäure ist aus WO-A-8606729 bekannt.A controlled chemical depolymerization of natural polysaccharides such as heparins by a radical reaction in the presence of a metal catalyst and a peroxide or a peracid is known from WO-A-8606729.
GB-A-978494 und Methods in Carbohydrate Chemistry, Vol. III (1963), 175-177, beschreiben die Behandlung linearer Polysaccharide mit Hypochlorit, jedoch hauptsächlich unter dem Gesichtspunkt der Oxidation des Polysaccharidmoleküls.GB-A-978494 and Methods in Carbohydrate Chemistry, Vol. III (1963), 175-177, describe the treatment of linear polysaccharides with hypochlorite, but mainly from the point of view of the oxidation of the polysaccharide molecule.
Andererseits beschreiben die von G.V. Buxton et al. in J. Chem. Soc., Faraday Trans. I, 958 (1972) die photolytische Zersetzung einer alkalischen wäßrigen Hypochlorition- Lösung.On the other hand, G.V. Buxton et al. in J. Chem. Soc., Faraday Trans. I, 958 (1972) describe the photolytic decomposition of an alkaline aqueous hypochlorite ion solution.
Das chemische Verfahren, das der Gegenstand der vorliegenden Erfindung ist, ist eine kontrollierte Depolymerisation von Heparin und anderen linearen Polysacchariden, die durch atomaren Sauerstoff als Mittel zur Spaltung der Glykosidbindung erhalten werden, worin der atomare Sauerstoff aus hypochloriger Säure in wäßriger Lösung bei einem pH-Wert zwischen 6 und 7 und bei einer Temperatur zwischen 25 und 100ºC erhalten wird.The chemical process which is the subject of the present invention is a controlled depolymerization of heparin and other linear polysaccharides obtained by using atomic oxygen as a glycosidic bond cleavage agent, wherein the atomic oxygen is obtained from hypochlorous acid in aqueous solution at a pH between 6 and 7 and at a temperature between 25 and 100°C.
Die Vorteile des Verfahrens der Erfindung sind zum Beispiel leichtes Stoppen der Reaktion bei jedem im voraus gewählten Molekulargewicht, hohe Ausbeuten, das Fehlen von Nebenreaktionen und das Fehlen anorganischer Katalysatoren (Schwermetalle). Ein weiterer Vorteil ist, daß, solange die Reaktion fortschreitet, keine Dunkelfärbung auftritt. Es wird daher ein weißes klares Produkt erhalten, und folglich werden komplizierte Bleichverfahren vermieden.The advantages of the process of the invention are, for example, easy stopping of the reaction at any preselected molecular weight, high yields, the absence of side reactions and the absence of inorganic catalysts (heavy metals). A further advantage is that as long as the reaction proceeds, no darkening occurs. A white clear product is therefore obtained and consequently complicated bleaching processes are avoided.
Das Verfahren der Erfindung, das die obigen Vorteile ermöglicht, wird unter bestimmten Reaktionsbedingungen durchgeführt, die wie folgt zusammengefaßt werden können:The process of the invention, which enables the above advantages, is carried out under certain reaction conditions, which can be summarized as follows:
1) Die wäßrige Lösung wird bei einem pH-Wert zwischen 6 und 7 gepuffert, um den PH-Wert während des gesamten Verfahrens konstant zu halten. Bei einem PH-Wert unter 4 oder im Falle der Erniedrigung des PH-Wertes unter 4 treten unerwünschte Nebenreaktionen aufgrund von Chlorfreisetzung aus hypochloriger Säure auf. Die gewünschte Reaktion (pH-Wert 6 bis 7) ist: HCl0 T HCl + 0. Andererseits ist die unerwünschte Reaktion (pH-Wert unter 4), die Chlorierung hervorruft: HCl0 + HCl T Cl&sub2; + H&sub2;.1) The aqueous solution is buffered at a pH between 6 and 7 to keep the pH constant throughout the process. At a pH below 4 or in case of a decrease of the pH below 4, undesirable side reactions occur due to chlorine release from hypochlorous acid. The desired reaction (pH 6 to 7) is: HCl0 → HCl + 0. On the other hand, the undesirable reaction (pH below 4) causing chlorination is: HCl0 + HCl → Cl₂ + H₂.
2) Die Reaktion wird bei einem Molverhältnis von HCl0/Polysaccharid 10:1 oder höher durchgeführt. Die gesamte veranschlagte Menge der HCl0-Lösung wird zu der Lösung der Polysaccharide zugegeben.2) The reaction is carried out at a molar ratio of HCl0/polysaccharide 10:1 or higher. The total estimated amount of HCl0 solution is added to the solution of polysaccharides.
3) Die Reaktion kann sofort bei einer im voraus gewählten Konzentration durch die Zugabe eines Reduktionsmittels, wie zum Beispiel Natriumhydrogensulfit NaHSO&sub3;, gestoppt werden, um überschüssiges HCl0 zu zerstören.3) The reaction can be stopped immediately at a preselected concentration by the addition of a reducing agent such as sodium hydrogen sulfite NaHSO3 to destroy excess HCl0.
4) Das depolymerisierte Polysaccharid wird durch Zugabe eines mit Wasser mischbaren Lösungsmittels, insbesondere von zwei Volumenteilen eines mit Wasser mischbaren Lösungsmittels wie Ethanol, Methanol oder Aceton gefällt, dann gesammelt und mit reinem Lösungsmittel gewaschen und unter Vakuum getrocknet.4) The depolymerized polysaccharide is precipitated by adding a water-miscible solvent, in particular two volumes of a water-miscible solvent such as ethanol, methanol or acetone, then collected and washed with pure solvent and dried under vacuum.
5) Der Betriebstemperaturbereich liegt zwischen 25ºC und 100ºC. Ein Temperaturbereich zwischen 50ºC und 80ºC wird üblicherweise verwendet, um die Reaktion zu beschleunigen und Rückflußkochen zu vermeiden.5) The operating temperature range is between 25ºC and 100ºC. A temperature range between 50ºC and 80ºC is commonly used to To accelerate the reaction and avoid reflux.
Die Temperatur der HCl0-Lösung vor der Zugabe der Polysaccharid-Lösung muß unter +5ºC gehalten werden.The temperature of the HCl0 solution before adding the polysaccharide solution must be kept below +5ºC.
6) Die Teil- und Endbestimmungen des mittleren Molekulargewichts können durch Viskosimetrie gemäß E.A. Johnson und B. Mulloy - Carbo hydr. Re. 51, 119 (1976) - gemacht werden.6) The partial and final determinations of the average molecular weight can be made by viscometry according to E.A. Johnson and B. Mulloy - Carbo hydr. Re. 51, 119 (1976).
Depolymerisierte Heparine, die mit therapeutisch nützlichen Eigenschaften ausgestattet sind und durch das oben beschriebene Verfahren erhalten werden, ebenso wie pharmazeutische Zusammensetzungen, die diese depolymerisierten Heparine enthalten, sind ein weiterer Gegenstand der Erfindung.Depolymerized heparins endowed with therapeutically useful properties and obtained by the process described above, as well as pharmaceutical compositions containing these depolymerized heparins, are a further subject of the invention.
Das folgende Beispiel, nicht beschränkend für das Verfahren, beschreibt die Art des Vorgehens:The following example, which is not limitative of the procedure, describes the way in which the procedure should be carried out:
15 g Heparin-Natrium USP (United States Pharmacopoeia), =-12.000 Daltons (bestimmt durch Viskosimetrie) wurden in 80 ml 0,1m Citrat/Phosphat-Puffer, pH = 6,0, gelöst. 20 ml 0,1m Natriumhypochlorit wurden getrennt hergestellt, bei 6 mit HCl stabilisiert und bei 0ºC aufbewahrt. Die Heparin-Lösung wurde zu dieser Lösung zugegeben. Auf diese Weise umfaßte die Endlösung 0,01 Mol Heparin und 0,2 Mol HCl0. Sie wurde auf 80ºC erhitzt, und alle 10 Min. wurde eine 10-ml-Probe isoliert. Sofort wurde 1 ml einer 2m NaHSO&sub3;-Lösung zu jeder Probe zugegeben und dann 20 ml Ethanol. Der Niederschlag wurde gesammelt, dreimal mit jeweils 5 ml Ethanol gewaschen und unter Vakuum getrocknet.15 g of heparin sodium USP (United States Pharmacopoeia), =-12,000 daltons (determined by viscometry) was dissolved in 80 ml of 0.1m citrate/phosphate buffer, pH = 6.0. 20 ml of 0.1m sodium hypochlorite was prepared separately, stabilized at 6 with HCl and stored at 0ºC. The heparin solution was added to this solution. Thus, the final solution comprised 0.01 mol heparin and 0.2 mol HCl0. It was heated to 80ºC and a 10 ml sample was isolated every 10 min. Immediately 1 ml of a 2m NaHSO₃ solution was added to each sample and then 20 ml ethanol. The precipitate was collected, washed three times with 5 ml of ethanol each and dried under vacuum.
Das wurde für jede Probe durch Viskosimetrie bestimmt. Die Ergebnisse sind in Fig. 1 wiedergegeben.This was determined for each sample by viscometry. The results are shown in Fig. 1.
Claims (6)
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
AR86305985A AR243204A1 (en) | 1986-11-21 | 1986-11-21 | A method for the chemical depolymerisation of polysaccharides. |
Publications (2)
Publication Number | Publication Date |
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DE3785726D1 DE3785726D1 (en) | 1993-06-09 |
DE3785726T2 true DE3785726T2 (en) | 1993-09-23 |
Family
ID=3478456
Family Applications (2)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
DE8787116225T Expired - Fee Related DE3785726T2 (en) | 1986-11-21 | 1987-11-04 | METHOD FOR CONTROLLED DEPOLYMERIZATION OF POLYSACCHARIDES. |
DE198787116225T Pending DE268885T1 (en) | 1986-11-21 | 1987-11-04 | METHOD FOR CONTROLLED DEPOLYMERIZATION OF POLYSACCHARIDES. |
Family Applications After (1)
Application Number | Title | Priority Date | Filing Date |
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DE198787116225T Pending DE268885T1 (en) | 1986-11-21 | 1987-11-04 | METHOD FOR CONTROLLED DEPOLYMERIZATION OF POLYSACCHARIDES. |
Country Status (8)
Country | Link |
---|---|
US (1) | US4977250A (en) |
EP (1) | EP0268885B1 (en) |
JP (1) | JPS63191801A (en) |
AR (1) | AR243204A1 (en) |
AT (1) | ATE89009T1 (en) |
DE (2) | DE3785726T2 (en) |
ES (1) | ES2054643T3 (en) |
IL (1) | IL84532A (en) |
Families Citing this family (18)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
AR243204A1 (en) * | 1986-11-21 | 1993-07-30 | Ajorca Sa | A method for the chemical depolymerisation of polysaccharides. |
US5280016A (en) * | 1991-03-29 | 1994-01-18 | Glycomed Incorporated | Non-anticoagulant heparin derivatives |
ES2049561B1 (en) * | 1991-04-27 | 1994-12-16 | Andromaco Lab | PROCEDURE FOR OBTAINING POLYMERS WITH ACTIVITY ON THE HEMATOPOYETIC SYSTEM. |
DE4138754C2 (en) * | 1991-11-26 | 1998-10-22 | Bayer Ag | Polycarbonate laundry |
JP2996559B2 (en) * | 1992-01-29 | 2000-01-11 | 本田技研工業株式会社 | Electric vehicle charging status display system |
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DK350078A (en) * | 1977-08-08 | 1979-02-09 | Choay Sa | PROCEDURE FOR PURIFICATION OF HEPARIN AND HEPARIN SALTS |
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US4446314A (en) * | 1980-09-30 | 1984-05-01 | Cutter Laboratories, Inc. | Fractionation of heparin |
FR2538404B1 (en) * | 1982-12-28 | 1985-08-23 | Anic Spa | |
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IT1214609B (en) * | 1985-05-17 | 1990-01-18 | Opocrin Spa | HEXOSAMINOGLICANS DEPOLYMERIZED SULPHATES FOR ANTI-THROMBOTIC, FIBRINOLITHIC, ANTI-INFLAMMATORY ACTIVITIES, THEIR PREPARATION PROCEDURE AND PHARMACEUTICAL COMPOSITIONS CONTAINING THEM. |
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US4767463A (en) * | 1987-04-15 | 1988-08-30 | Union Carbide Corporation | Glycosaminoglycan and cationic polymer combinations |
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1986
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- 1987-11-04 EP EP87116225A patent/EP0268885B1/en not_active Expired - Lifetime
- 1987-11-04 DE DE198787116225T patent/DE268885T1/en active Pending
- 1987-11-19 US US07/122,466 patent/US4977250A/en not_active Expired - Fee Related
- 1987-11-19 IL IL84532A patent/IL84532A/en not_active IP Right Cessation
- 1987-11-19 JP JP62293060A patent/JPS63191801A/en active Pending
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IL84532A0 (en) | 1988-04-29 |
IL84532A (en) | 1992-08-18 |
DE3785726D1 (en) | 1993-06-09 |
AR243204A1 (en) | 1993-07-30 |
ES2054643T3 (en) | 1994-08-16 |
ATE89009T1 (en) | 1993-05-15 |
DE268885T1 (en) | 1988-11-24 |
EP0268885B1 (en) | 1993-05-05 |
JPS63191801A (en) | 1988-08-09 |
EP0268885A2 (en) | 1988-06-01 |
US4977250A (en) | 1990-12-11 |
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